Over half of all heart failure (HF) diagnoses are HF with preserved ejection fraction (HFpEF), a complex syndrome defined by symptoms of HF but a `normal' left ventricular EF. HFpEF is characterized by diastolic dysfunction, impaired filling, hypertrophy, and elevated natriuretic peptides. Presently, there are no treatments that improve survival in patients with HFpEF. Pathologic remodeling in HFpEF includes interstitial fibrosis and microvascular rarefaction, which contribute to diastolic stiffness, impaired ventricular filling, and exercise intolerance. Fibrosis results from activation of fibroblasts leading to increased extracellular matrix production, fibroblast proliferation, and fibroblast-to-myofibroblast transformation. Microvascular rarefaction is associated with inflammation and endothelial cell death, leading to impaired oxygen delivery, reducing systolic and diastolic reserve, and exacerbating exercise intolerance. We found that ?3-polyunsaturated fatty acids (?3- PUFA) inhibited myofibroblast transformation and prevented interstitial fibrosis and diastolic dysfunction in a mouse model that partially resembles HFpEF (transverse aortic constriction, TAC). Further, we found that blood levels of eicosapentaenoic acid (EPA) correlated with prevention of interstitial fibrosis, but EPA was not incorporated into fibroblast membranes in vivo, a traditional mechanism of action. Rather, we found that free fatty acid receptor 4 (FFAR4), a G-protein coupled receptor for long-chain fatty acids, was sufficient and required to prevent myofibroblast transformation in cardiac fibroblasts. Other groups reported that ?3-PUFAs induce angiogenesis in hind-limb ischemia, and EPA activates FFAR4 to induce VEGF-A expression. Thus, EPA might have a dual benefit in HFpEF by preventing fibrosis and microvascular rarefaction. In humans, ?3- PUFAs reduce sudden death in coronary heart disease, but questions of efficacy remain. However, failure to achieve adequate ?3-PUFA levels, or a therapeutic index, likely explains prior negative results. Thus, we hypothesize that sufficient dietary uptake of EPA (therapeutic level) and activation of FFAR4 prevents fibrosis, microvascular rarefaction, and diastolic dysfunction in HFpEF. To test this we will: 1) Determine if erythrocyte EPA concentration (EPA-index) correlates with prevention of fibrosis, microvascular rarefaction, and diastolic dysfunction following TAC. We will also target FFAR4 with cpdA, a novel FFAR4 ligand, as a potential therapeutic approach to reverse remodeling following TAC. 2) Determine if high plasma levels of EPA prevent progression of left ventricular hypertrophy and fibrosis as well as HFpEF in humans using data from the Multi- Ethnic Study of Atherosclerosis trial (MESA). 3) Use fibroblast-specific FFAR4 knockout mice to determine if FFAR4 is required for EPA mediated prevention of fibrosis, microvascular rarefaction, and diastolic dysfunction following TAC. The impact will be to: 1) Identify novel cardioprotective functions of EPA, 2) Define a therapeutic index for EPA-mediated cardioprotection, 3) Provide evidence for the clinical efficacy of EPA/ FFAR4 to prevent HFpEF, and 4) Delineate a novel mechanism of action for EPA, activation of FFAR4.